Promoting Effect of Soluble Polysaccharides Extracted from Ulva spp. on Zea mays L. Growth

Seaweeds can play a vital role in plant growth promotion. Two concentrations (5 and 10 mg/mL) of soluble polysaccharides extracted from the green macroalgae Ulva fasciata and Ulva lactuca were tested on Zea mays L. The carbohydrate and protein contents, and antioxidant activities (phenols, ascorbic, peroxidase, and catalase) were measured, as well as the protein banding patterns. The soluble polysaccharides at 5 mg/mL had the greatest effect on the base of all of the parameters. The highest effects of soluble polysaccharides on the Zea mays were 38.453, 96.76, 4, 835, 1.658, 7.462, and 38615.19, mg/mL for carbohydrates, proteins, phenol, µg ascorbic/mL, mg peroxidase/g dry tissue, and units/g tissue of catalase, respectively. The total number of protein bands (as determined by SDS PAGE) was not changed, but the density of the bands was correlated to the treatments. The highest band density and promoting effect were correlated to 5 mg/mL soluble polysaccharide treatments extracted from Ulva fasciata in Zea mays, which can be used as a biofertilizer.


Introduction
The third most vital cereal crop in the world is maize (Zea mays L.), which comes after wheat, followed by rice [1]. Every portion of the maize plant has an efficient value, such as the grain, which is consumed in the production cooking oil; and the leaves, stalks, tassels, and cob, all of which can be consumed in animal feed and also used in the production of bioethanol [2,3]. The excess use of chemical synthetic fertilizers has led to the contamination of the soil and water basins, destroying micro-organisms and destroying friendly insects, which renders the crop more susceptible to infections and reduces soil fertility [4]. Biofertilizers are of the highest significance as a complement to synthetic chemical fertilizers in enhancing the nutrient materials provided to cereal crops due to the great nutrient turn-over in the cereal-yields system [5]. Algae could be a possible

Algae
U. facciata and U. lactuca were gathered from the shallow water on the beach of the Mediterranean Sea at Abo Quire coast, Alexandria. The collected algae were cleaned using tap water and dried in an electric oven at 60 • C until they reached a constant weight.

Extraction of Water-Soluble Polysaccharides
The sulphated polysaccharide was extracted by the procedure detailed by Bourgougnon et al. [22]. Approximately 1500 mL of distilled water was added to 5 g each U. fasciata and U. lactuca, which was then heated to 80 • C for 4 h with magnetic stirring; after that, the suspension was cooled to 5 • C in a refrigerator, and filtered using Whatman no. 1 filter paper. Two volumes of absolute ethanol were added to the filtrate, with stirring. The precipitate was recovered, cleaned with ethanol, dehydrated with diethyl ether, and dried overnight at 50 • C.

Physical Characterizations Thermal Characteristics of the Extracted Polysaccharides
The thermal stability of the extracted polysaccharide was investigated using thermogravimetric analysis (TGA) with a Shimadzu D-50 thermo-analyzer (Japan). The polysac-charide sample was heated at a rate of 15 • C/min in a dry nitrogen environment, and the thermal interference material was sintered alumina (α-alumina). The temperature was then measured with the aid of a Pt/Rh (10%) thermocouple.
The thermal stability of the extracted polysaccharide was investigated using thermogravimetric analysis (TGA) with a Shimadzu D-50 thermo-analyzer (Japan).

Chemical Characterizations
High-Performance Liquid Chromatography (HPLC)-Refractive Index Detector The monosaccharide analysis of the extracts of the decomposed algae was performed by HPLC (Agilent 1100 HPLC system) using a Hypersil ASP-2 column (4.6 × 250 mm) with a mobile phase of acetonitrile-water (80:20) and a flow rate of 0.4 mL min −1 . The column and optical unit were both heated to 35 and 40 • C, respectively. The injection volume of each integrated monosaccharide standard and the sample hydrolyses was 10µL.

Biological Activity
Investigation of the Promoting Growth Efficiency of the Extracted Water-Soluble Polysaccharides

Plant Growth Analysis
The Ministry of Agriculture, Field Crop Institute, Agriculture Research Center, Giza, Egypt, provided seeds of Zea mays single cross 10. They were tested for size and shape uniformity before being surface-sterilized in 0.01 percent HgCl 2 for 3 min [23]. They were then cleaned carefully with running distilled water. Following that, the seeds were soaked in soluble polysaccharides (10 mg mL -1 , 5 mg mL -1 ) or water (control) for 4 h at 25 • C. The seeds were rinsed with sterile water for 15 min.

Determination of Germination
The germination experiment was conducted as follows: The same number of seeds (10 seeds) were germinated in Petri dishes (11 cm) lined with two moist sheets of filter paper (Whatman no.1) and incubated in the dark at 25 • C for 5 days in order to determine the germination criteria, such as the percentage of seed germination, the radical length (the samples were ten replicates), and the amylase or protease activity (in triplicate). Another series of soaked seeds was grown in plastic pots (1 Kg sand soil capacity) and incubated under continuous illumination (72 µmol photon m −2 s −1 ) at 25 • C for 14 days; the sampling was performed on the seventh day and at the end of the experimental period (14 days). The growth criteria of the seedlings-such as shoot height and root length, leaf area, fresh and dry weight, and leaf area (ten replicates for each), as well as the photosynthetic pigment, carbohydrate, and protein content (triplicate sampling)-were determined. In addition, the antioxidant activities (triplicate sampling) were also determined.

Phytochemical Contents
The total carbohydrate contents were assessed with the phenol-sulphuric acid method, using glucose as the standard [24]. The protein content was investigated according to the method of Lowry et al. [25], using crystalline bovine serum albumin as the standard. The total phenol content was assessed by the Folin-Ciocalteu method [26].

Total Soluble Proteins Banding Pattern for the Tested Crop Plant
The discontinuous polyacrylamide gel electrophoresis (SDS-PAGE) was exhausted for the protein analysis, allowing the method adopted by Stegman et al. [27] and Hemeida [28].

Antioxidants Total Ascorbic Acid Content
The total ascorbic acid content was assessed as indicated by Chen and Asada [29]. The CAT activity was assayed following the method of Kang and Saltveit [30].

Peroxidase
The activity was assayed following the method of Racusen and Foote [31].

Amylase Assay
The activity was assayed following the method of Monroe and Preiss [32].

Protease Assay
The protease was assayed according to Anson [33].

Statistical Analysis
The statistical analysis was conveyed by using ANOVA, analysis of variance (version 19) for comparison between the treatments according to Sokal and Rohlf [34]. Significant differences between the means of the parameters were investigated using Duncan's multiple range tests, with a probability of ≤0.05. All of the previously mentioned measurable investigations were focused using SPSS software.

Results and Discussion
The chemical compositions of the U. lactuca and U. fasciata are shown in Table 1. The main component of U. fasciata and U. lactuca is the carbohydrate fraction, which occupies about 58.5 and 56.8 % on a dry-weight basis, followed by a protein which represents about 27.7 and 25.9 % of the dry weight. The lipid content didn't exceed 1% in the two analyzed algal dry tissues, while the highest value of ash content was recorded for U. lactuca. Pauline et al. [8] and Taboada et al. [9] showed that Ulva sp. contains high quantities of beneficial-quality protein, carbohydrates, vitamins, and minerals. Research has displayed increased attention to the applications of polysaccharides due to their unique chemical and physical properties [35]. Ulvan is a form of sulphated heteropolysaccharide isolated from the cell walls of Ulva sp. marine green alga, the composition of which has been widely deliberated [36,37]. Ulvan is a sulphated heteropolysaccharide derived from the cell wall of Ulva sp. green seaweeds, the composition of which has been fiercely debated; it is a major biopolymeric fraction of the cell wall that is responsible for preserving the cell's osmolar stability and defense [38]. Ulva. sp. has a high protein content, similar to legumes and grains, especially amaranth and soy [39]. The results revealed that Ulva sp has a low amount of lipids, and the ash content is much greater than that present in the higher plant; these results are in agreement with [40].  [41]. According to the present thermogravimetric results, the extracted polysaccharides from the green algae U. lactuca and U. fasciata are thermally stable until about 169 and 245 • C, respectively. These values are in agreement with those recorded by Alves et al. [42]. Table 2. TGA analysis of the extracted polysaccharides from U. fasciata and U. lactuca.

Chemical Characterization of the Extracted Polysaccharide
The chemical compositions of the polysaccharides extracted from U. lactuca and U. fasciata is given in Figure 1 and Table 3. There were various types of sugars in the samples, such as xylose, glucose, rhamnose (in U. lactuca only), galactose, and arabinose. Each polysaccharide has a different composition than the others. Rhamnose, glucose, and galactose were the most abundant sugars. Ulvan is a form of sulphated heteropolysaccharide obtained from the cell walls of Ulva sp. green seaweeds, the composition of which has been extensively discussed [36,37]. Glucose is produced from hemicellulose and cellulose. The absence of fructose in the algal biomasses proved the absence of sucrose. The presence of rhamnose may be due to the existence of xylorhamnoglycuronane in this seaweed [36,37]. The variations in the sugar proportions cannot be attributed to the species or to the ecophysiological conditions [43]. Investigating the biological activity, Figure 2 shows the changes in the germination percentage of the seeds treated with the extracted polysaccharides. The germination percentage increased progressively throughout the germination period (5 days) of all of the treatments. Compared to the control level, highly significant (p value = 0.000) increases in the germination percentage of Zea mays were found in the following descending order: U. fasciata. polysacch. 5 mg/mL > U. fasciata. polysacch. 10 mg/mL > U. lactuca. polysacch., 5 mg/mL > U. lactuca. polysacch., 10 mg/mL priming solution agent. Due to the high levels of organic matter, micro and macro elements, vitamins, and fatty acids, as well as being rich in growth regulators, liquid fertilizers originating from seaweeds are better than chemical fertilizers [44]. In the life cycle of flowering plants, seed germination is crucial. The germination process starts with the ingestion of dry seeds and ends with the embryo's radical protrusion through the covering tissues [45]. Seaweed products have been shown to improve seed germination, improve nutrient absorption, enhance frost resistance, and help plants to resist phytopathological fungi and insect pests [46]. Furthermore, it was discovered that dilute extracts were more efficient than concentrated extracts [47]. Paulert et al. [48] found that using 10 mg/mL ulvan solute extracted from Ulva fasciata as a priming agent for Phaseolus vulgaris seeds increased the seed germination by about 24%. The present results follow those of Ramarajan et al. [49], who investigated the effect of seaweed extracts of Ulva lactuca and Sargassum wightii on bean seeds. They recorded that priming cluster bean seeds with 1% and 2% seaweed extracts enhanced the seed germination over the control level. The higher promotion effect was recorded at the lower concentration (1%) of seaweeds, and the ameliorating effect was attributed to Ulva rather than the Sargassum extract. This ameliorating effect may be attributed to the promotion effect of the water-soluble polysaccharide under investigation on a growthpromoting substance. The treated seeds with a green seaweed suspension had a significant stimulatory effect on germination, as they reduced the mean germination time at the optimum temperature in tomato and pepper seeds [50]. The active ingredients in seaweed extracts are efficient at low concentrations. Seaweed extracts enhance seed germination in several species, such as Cajanus cajan [51], Trigonella foenum [52], and Capsicum annum [53]. Seaweed extracts were found to improve seed germination in ornamental plants, tobacco, peas, and cotton at lower doses [54].
Seaweed liquid fertilizer (SLF) promoted the level of seed germination in green chilies and turnip, and it was found that minor concentrations of SLF promoted the germination percentage better than the superior concentration [13]. It was indicated that the promoting effect of seaweed extracts may originate from the higher level of seed moisture around the seeds [55]. In the present work, soluble polysaccharide priming agents (5 and 10 mg/mL of U. fasciata and U. lactuca polysaccharide) induced significant promotion effects on seed germination and seedling growth. It is noteworthy that these concentration doses lead to phytotoxicity. Cluz et al. [56] previously reported that the polysaccharide from Ulva spp. was non-phytotoxic.

Radical Length
Figures 3 and 4 indicate the lengths of the radicles of the control and the treated seedlings of Zea mays appeared to increase progressively throughout the germination period (5 days). The Zea mays seedlings, primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in radicle length, whereas significant increases were found upon priming with U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL. Kavipriy et al. [57] reported that seaweed extracts attained from seaweeds such as Ulva lactuca, Caulerpa scalpelliformis, Sargassum plagiophyllum, Turbinaria conoides, Padina tetrastromatica, and Dictyota dichotama strongly induce the seed germination and growth parameters of Vigna radiata. Among the four marine algae (Ulva lactuca, Jania rubens, Laurencia obtusa and Sargassum vulgares) which were tested, Ulva lactuca was the best alga for effectiveness in the promotion of banana plants' growth [58].

Morphological Criteria
The data presented in Figures 5 and 6 indicate the promoting action on the morphological criteria of the tested plants compared with the control values. The Zea mays seedlings primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in shoot and root length, whereas a significant increase was found upon priming with both U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL.

Fresh Weight
The data presented in Figure 7 shows that the fresh weights of the treated seedlings of Zea mays increased progressively throughout the experimental period (14 days). Highly significant increases in fresh weight were recorded in the following descending order: U.f. polysacch. 5 mg/mL > U.f. polysacch. 10 mg/mL > U.l. polysacch. 5 mg/mL > U.l. polysacch. 10 mg/mL.

Radical Length
Figures 3 and 4 indicate the lengths of the radicles of the control and the treated seedlings of Zea mays appeared to increase progressively throughout the germination period (5 days). The Zea mays seedlings, primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in radicle length, whereas significant increases were found upon priming with U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL. Kavipriy et al. [57] reported that seaweed extracts attained from seaweeds such as Ulva lactuca, Caulerpa scalpelliformis, Sargassum plagiophyllum, Turbinaria conoides, Padina tetrastromatica, and Dictyota dichotama strongly induce the seed germination and growth parameters of Vigna radiata. Among the four marine algae (Ulva lactuca, Jania rubens, Laurencia obtusa and Sargassum vulgares) which were tested, Ulva lactuca was the best alga for effectiveness in the promotion of banana plants' growth [58].

Radical Length
Figures 3 and 4 indicate the lengths of the radicles of the control and the treated seedlings of Zea mays appeared to increase progressively throughout the germination period (5 days). The Zea mays seedlings, primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in radicle length, whereas significant increases were found upon priming with U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL. Kavipriy et al. [57] reported that seaweed extracts attained from seaweeds such as Ulva lactuca, Caulerpa scalpelliformis, Sargassum plagiophyllum, Turbinaria conoides, Padina tetrastromatica, and Dictyota dichotama strongly induce the seed germination and growth parameters of Vigna radiata. Among the four marine algae (Ulva lactuca, Jania rubens, Laurencia obtusa and Sargassum vulgares) which were tested, Ulva lactuca was the best alga for effectiveness in the promotion of banana plants' growth [58].

Morphological Criteria
The data presented in Figures 5 and 6 indicate the promoting action on the morphological criteria of the tested plants compared with the control values. The Zea mays seedlings primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in shoot and root length, whereas a significant increase was found upon priming with both U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL.

Morphological Criteria
The data presented in Figures 5 and 6 indicate the promoting action on the morphological criteria of the tested plants compared with the control values. The Zea mays seedlings primed with U.f. polysacch. 5 mg/mL and U.f. polysacch. 10 mg/mL, as compared with the control values, induced highly significant increases in shoot and root length, whereas a significant increase was found upon priming with both U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL.

Fresh Weight
The data presented in Figure 7 shows that the fresh weights of the treated seedlings of Zea mays increased progressively throughout the experimental period (14 days). Highly significant increases in fresh weight were recorded in the following descending order: U.f. polysacch. 5 mg/mL > U.f. polysacch. 10 mg/mL > U.l. polysacch. 5 mg/mL > U.l. polysacch. 10 mg/mL.

Fresh Weight
The data presented in Figure 7 shows that the fresh weights of the treated seedlings of Zea mays increased progressively throughout the experimental period (14 days). Highly significant increases in fresh weight were recorded in the following descending order: U.f. polysacch. 5 mg/mL > U.f. polysacch. 10 mg/mL > U.l. polysacch. 5 mg/mL > U.l. polysacch. 10 mg/mL.   Figure 8 shows the changes in the dry weight of the treated seeds of Zea mays. There appears to be a general progressive increase in the dry weight of the treated seeds of Zea mays throughout the germination period. All of the polysaccharide treatments induced significant increases in the dry weight of the treated seedlings, giving the following trend: U.f. polysacch. 5mg/mL > U.f. polysacch. 10mg/mL > U.l. polysacch. 5mg/mL > U.l. polysacch. 10mg/mL compared with the control values. Lower doses of seaweed liquid fertilizer (SLF) of Chaetomorpha linum (30%) were more effective regarding the growth parameters than higher doses (100%) when Cajanus cajan seeds were rinsed with it, along with lower concentrations of SLF extracted from Stoechospermum marginatum [59]. Similar effects were obtained with lower concentrations of seaweeds on the promotion of plants such as Ascophyllum and Laminaria, and accelerated maize growth [60]. Padina induced the maximum growth in Cajanus cajan [61], Stoechosperum marginatum, Hypnea musciformis, Spatoglossum asperum, and Sargassum with the growth of green chilies, pineapples, and turnips [62]. Selvam et al. [63] recorded that Vigna mungo seeds soaked with lower concentrations (1, 2.5, and 5 %) of seaweed extract prepared from the marine green algae Ulva reticulate showed higher rates of germination and enhanced shoot and root length, while the higher concentrations (7.5 and 10%) of the extract inhibited the germination. significant increases in the dry weight of the treated seedlings, giving the following trend: U.f. polysacch. 5mg/mL > U.f. polysacch. 10mg/mL > U.l. polysacch. 5mg/mL > U.l. polysacch. 10mg/mL compared with the control values. Lower doses of seaweed liquid fertilizer (SLF) of Chaetomorpha linum (30%) were more effective regarding the growth parameters than higher doses (100%) when Cajanus cajan seeds were rinsed with it, along with lower concentrations of SLF extracted from Stoechospermum marginatum [59]. Similar effects were obtained with lower concentrations of seaweeds on the promotion of plants such as Ascophyllum and Laminaria, and accelerated maize growth [60]. Padina induced the maximum growth in Cajanus cajan [61], Stoechosperum marginatum, Hypnea musciformis, Spatoglossum asperum, and Sargassum with the growth of green chilies, pineapples, and turnips [62]. Selvam et al. [63] recorded that Vigna mungo seeds soaked with lower concentrations (1, 2.5, and 5 %) of seaweed extract prepared from the marine green algae Ulva reticulate showed higher rates of germination and enhanced shoot and root length, while the higher concentrations (7.5 and 10%) of the extract inhibited the germination.

Changes in the Photosynthetic Pigments
The results regarding the photosynthetic pigments are presented in Figure 9, and make it clear that the chlorophylls a and b, carotenoids, and total pigments at the vegetative stage of the treated Zea mays appeared to increase significantly in the following trend: U.f. polysacch., 5mg/mL, U.f., polysacch. 10 mg/mL, U.l. polysacch. 5 mg/mL, 10 mg/mL U.l., polysacch., respectively.

Changes in the Photosynthetic Pigments
The results regarding the photosynthetic pigments are presented in Figure 9, and make it clear that the chlorophylls a and b, carotenoids, and total pigments at the vegetative stage of the treated Zea mays appeared to increase significantly in the following trend: U.f. polysacch., 5mg/mL, U.f., polysacch. 10 mg/mL, U.l. polysacch. 5 mg/mL, 10 mg/mL U.l., polysacch., respectively.
The increased level of chlorophyll observed in this study was supported by the findings of Blunden and Wildgoose [64], who reported a marked enhancement in the lateral root growth of potato plants as a result of the effect of seaweed extract. Stirk et al. [65] suggested that the secondary metabolites present in the polysaccharide extracts may stimulate growth by increasing the photosynthetic area of the treated seedlings.
Moreover, seaweed extracts such as brown algae have been added to many crops as promoters, with physiological promotions such as developed nutrient mobilization and partitioning, the advance of a vigorous root system, and improved chlorophyll content and leaf area. Christobel [66] deliberated the effects of seaweed liquid fertilizer (SLF) of Sargassum wightii on the germination, seedling growth, and phytochemical content of green gram (Phaseolus aureus). He found that a low level of seaweed liquid fertilizer of Sargassum wightii (1%) enhances the germination percentage and seedling growth, viz., root length, shoot length, leaves, chlorophyll content (chlorophyll a and b, and total chlorophyll), and enzyme activity. At higher levels (10 and 25%), the effect was inhibitive. The increased level of chlorophyll observed in this study was supported by the findings of Blunden and Wildgoose [64], who reported a marked enhancement in the lateral root growth of potato plants as a result of the effect of seaweed extract. Stirk et al. [65] suggested that the secondary metabolites present in the polysaccharide extracts may stimulate growth by increasing the photosynthetic area of the treated seedlings.
Moreover, seaweed extracts such as brown algae have been added to many crops as promoters, with physiological promotions such as developed nutrient mobilization and partitioning, the advance of a vigorous root system, and improved chlorophyll content and leaf area. Christobel [66] deliberated the effects of seaweed liquid fertilizer (SLF) of Sargassum wightii on the germination, seedling growth, and phytochemical content of green gram (Phaseolus aureus). He found that a low level of seaweed liquid fertilizer of Sargassum wightii (1%) enhances the germination percentage and seedling growth, viz., root length, shoot length, leaves, chlorophyll content (chlorophyll a and b, and total chlorophyll), and enzyme activity. At higher levels (10 and 25%), the effect was inhibitive.
Thirumaran et al. [67] deliberated the influence of seaweed liquid fertilizer (SLF) on the growth and photosynthetic pigment of Cyamopsis tetrogonolaba. They found that the elevation of seed germination, growth and yield parameters, and photosynthetic pigment concentration were enhanced when treated with seaweed liquid fertilizer (SLF) at a 20% concentration, with or without chemical fertilizer. Francisca and Kalavathy [68] reported that aqueous extracts of Sargassum wightii and Ulva lactuca induced positive and significant responses in the growth and chlorophyll, protein, and carbohydrate contents of Zea mays plants. The increase in pigment production led to an improvement in the photosynthetic activity and carbohydrate content of the plant tissue. These findings could help to improve photosynthetic electron transport [69], pigment biosynthesis [70], and the inter- Thirumaran et al. [67] deliberated the influence of seaweed liquid fertilizer (SLF) on the growth and photosynthetic pigment of Cyamopsis tetrogonolaba. They found that the elevation of seed germination, growth and yield parameters, and photosynthetic pigment concentration were enhanced when treated with seaweed liquid fertilizer (SLF) at a 20% concentration, with or without chemical fertilizer. Francisca and Kalavathy [68] reported that aqueous extracts of Sargassum wightii and Ulva lactuca induced positive and significant responses in the growth and chlorophyll, protein, and carbohydrate contents of Zea mays plants. The increase in pigment production led to an improvement in the photosynthetic activity and carbohydrate content of the plant tissue. These findings could help to improve photosynthetic electron transport [69], pigment biosynthesis [70], and the interface with the thylakoid membrane surface [71]. Using seaweed extract as fertilizers increases the seed germination, seedling growth, and yield of the crop [72]. The micro-green alga Scenedesmus obliquus promoted banana plants' growth with regard to shoot and root length and weight [73].

Carbohydrate Contents
As compared with the control values, the perusal of the data presented in Figure 10 indicates that the highest productions of total carbohydrates and polysaccharides which were detected in the tested crop plant Zea mays with the treatments of U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, and U.l. polysacch. 5 mg/mL, were 38.453, 29.9 and 27.27 mg sugar/gm dry weight, while the lowest production of carbohydrates which was recorded in the tested crop plant with the treatments of U.l. polysacch. 10 mg/mL was 25.26 mg sugar/gm dry weight. Our results coincide with those of El-Sheekh and El-Saied, [74], who reported that the total soluble sugars were elevated in both the shoot and root of Vicia faba with treatments of Enteromorpha intestinalis, Cladophora dalmatica, Ulva lactuca, Corollina mediterranea, Pterocladia pinnata and Jania rubens extracts. Abdel-Hamid et al. [75] reported that the consortium of Chlorella vulgaris and Anabaena oryzae enhanced Jerusalem artichokes' growth, yield, and sugar contents.
were detected in the tested crop plant Zea mays with the treatments of U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, and U.l. polysacch. 5 mg/mL, were 38.453, 29.9 and 27.27 mg sugar/gm dry weight, while the lowest production of carbohydrates which was recorded in the tested crop plant with the treatments of U.l. polysacch. 10 mg/mL was 25.26 mg sugar/gm dry weight. Our results coincide with those of El-Sheekh and El-Saied, [74], who reported that the total soluble sugars were elevated in both the shoot and root of Vicia faba with treatments of Enteromorpha intestinalis, Cladophora dalmatica, Ulva lactuca, Corollina mediterranea, Pterocladia pinnata and Jania rubens extracts. Abdel-Hamid et al. [75] reported that the consortium of Chlorella vulgaris and Anabaena oryzae enhanced Jerusalem artichokes' growth, yield, and sugar contents.

Changes in Enzymatic Activities
The full data of the changes in the enzymatic activities of the growing seedlings, as affected by different concentrations of priming soluble polysaccharides, are presented in Table 4. Compared with the control values, soluble polysaccharide treatments of U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL induced significant increases in the amylase activity of Zea mays in seedlings. The protease activity of germinating seedlings of Zea mays appeared to increase in response to all of the soluble polysaccharide treatments. The changes in the catalase and peroxidase contents of the investigated seedlings of Zea mays affected by priming with different concentrations of soluble polysaccharides were 38615.19 units/g tissue and 7.462 mg/g dry tissue, respectively. The catalase and peroxidase contents of 14-day-old treated seedlings of Zea mays exhibited significant escalations above the control level in response to soaking with soluble polysaccharides of U. fasciata, at 5 and 10 mg/mL, and U.lactuca, at 5 and 10 mg/mL, respectively. The amylase and protease activities of germinating Zea mays seeds showed a significant stimulatory effect in response to the priming agents of Ulva spp. soluble polysaccharide. Kato-Noguchi and Macias [76] found that the percentage of germination was correlated with the activity of α-amylase in lettuce seeds. The breakdown of proteins in the Zea mays germinating seeds is accomplished by the activities of protease and peptidase [77]. In germination, α-amylase is thought to play a key role in the conversion of reserve carbohydrates to soluble sugars [78]. Therefore, the stimulation

Changes in Enzymatic Activities
The full data of the changes in the enzymatic activities of the growing seedlings, as affected by different concentrations of priming soluble polysaccharides, are presented in Table 4. Compared with the control values, soluble polysaccharide treatments of U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, U.l. polysacch. 5 mg/mL and U.l. polysacch. 10 mg/mL induced significant increases in the amylase activity of Zea mays in seedlings. The protease activity of germinating seedlings of Zea mays appeared to increase in response to all of the soluble polysaccharide treatments. The changes in the catalase and peroxidase contents of the investigated seedlings of Zea mays affected by priming with different concentrations of soluble polysaccharides were 38615.19 units/g tissue and 7.462 mg/g dry tissue, respectively. The catalase and peroxidase contents of 14-day-old treated seedlings of Zea mays exhibited significant escalations above the control level in response to soaking with soluble polysaccharides of U. fasciata, at 5 and 10 mg/mL, and U.lactuca, at 5 and 10 mg/mL, respectively. The amylase and protease activities of germinating Zea mays seeds showed a significant stimulatory effect in response to the priming agents of Ulva spp. soluble polysaccharide. Kato-Noguchi and Macias [76] found that the percentage of germination was correlated with the activity of α-amylase in lettuce seeds. The breakdown of proteins in the Zea mays germinating seeds is accomplished by the activities of protease and peptidase [77]. In germination, α-amylase is thought to play a key role in the conversion of reserve carbohydrates to soluble sugars [78]. Therefore, the stimulation of α-amylase is important in order to maintain an active respiratory metabolism, which permits the germination of plant seeds. The combinations of Turbinaria ornata and Ulva reticulate enhanced the germination and growth parameters of radish, green pea, and mung [79]. The amylase activity is calculated as the mg reducing sugars. min −1 ·g −1 fresh weight. The protease activity is calculated as the µg amino acids. min −1 ·g −1 fresh weight. The catalase content is given in units/g tissue, and the peroxidase content is given in mg/g dry tissue. Different letters denote the significant values among the treatments with each enzyme. Table 5 shows the changes in the phenol content and total ascorbic acid content of the various investigated plant seedlings of Zea mays, as affected by priming in different concentrations of soluble polysaccharides. The total phenol content and total ascorbic acid content of 14-day-old treated Zea mays seedlings were significantly higher than the control level in response to priming with U. fasciata at 5 and 10 mg/mL, and U. lactuca at 5 mg/mL, respectively. The major contents of phenol and the total ascorbic acid were 4.835 mg/g dry weight and 1.658 µg/g dry weight with 5 mg/mL U. fasciata, respectively. Table 5. Effect of seed priming with different concentrations of soluble polysaccharides on the ascorbic acid content (µg/g dry tissue) and total phenolic content (mg/g dry tissue) of Zea mays. Catalase plays an important role in an antioxidant system with a specific function. Catalase degrades H 2 O 2 (one of the very common ROS in plant tissues) into H 2 O and oxygen, where ASA serves as an electron donor to active H 2 O 2 degradation by ascorbate peroxidase (APX) [80]. Catalase (CAT) activity was accompanied by improved germination performance in sunflower seeds [81], and also in purple and coneflower [82], Arabidopsis [83], and tomato [84]. Ulvan increased the peroxidase activity in beans (Phaseolus vulgaris) [85]. The treatment of higher plants with Ulvan prompts plant resistance and decreases the impact and severity of fungal infections [86]. Ulvan activity may be related to the presence of sulphate besides the presence of uronic acid and rhamnose in its contents, acting through the jasmonic acid pathway [87]. Several types of sulphated polysaccharides have been shown to exhibit antioxidant activity. Among them, Ulvan isolated from Ulva pertusa was found to play a vital role as a free radical scavenger in vitro, and exhibited antioxidant activity for the inhibition of oxidative damage in living organisms [88].
Phenols are very essential plant contents due to their scavenging capability and their hydroxyl groups [89]. A recent study demonstrated that a major positive relationship between the total phenol content and antioxidant activity was obtained in several higher plants [90]. The antioxidant and phenol contents increased in banana plants when they were treated with algal combinations of Anabena oryza, Chlorella vulgaris and Scenedesmus obliquus [91]. Ascorbic acid (vitamin C) is a water-soluble antioxidant molecule that acts as a basic substance in the cyclic pathway for the enzymatic detoxification of hydrogen peroxide; it acts directly to neutralize superoxide radicals, singlet oxygen, or superoxide [92,93]. The ascorbic acid (ASA) content of Zea mays seeds treated with the water-soluble polysaccharides of either U. fasciata or U. lactuca had significantly higher values than those of the control, as recorded in Table 5.

Protein Content
As the data presented in Figure 11 indicate, U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, and U.l. polysacch. 5 mg/mL induced the highest production of protein content, and U.l. polysacch. 10 mg/mL induced the lowest of the different selected plant seedlings of Zea mays; the protein contents were 96.76, 85.49, 74.54 and 69.75, respectively. These results follow those of Christobel [66] and Asir and Saravanababu [94], who pointed out that there was an increase in the protein content at the lower concentrations of seaweed extract-treated plants. In the present study, the observed significant increases in the amylase and protease activities of both tested plant seedlings in response to the effect of the priming agents are in good accord with the rise in growth, as well as the carbohydrate and protein composition. The results show that the low concentrations of both U.f. polysacch and U.l. polysacch are more effective with regard to growth parameters, antioxdant activities and protein contents; this may be due to the high concentrations of polysaccharide having more viscosity than the low concentrations. These results are comparable to those obtained by Siddhanat et al. [95], who reported that increasing the concentration of Ulva polysaccharide solutions lead to an increase in viscosity. The lower viscosity of chitosan improved the stability of the soil aggregates by 100 to 200 times after just one cycle of wetting-drying [96].
obliquus [91]. Ascorbic acid (vitamin C) is a water-soluble antioxidant molecule that acts as a basic substance in the cyclic pathway for the enzymatic detoxification of hydrogen peroxide; it acts directly to neutralize superoxide radicals, singlet oxygen, or superoxide [92,93]. The ascorbic acid (ASA) content of Zea mays seeds treated with the water-soluble polysaccharides of either U. fasciata or U. lactuca had significantly higher values than those of the control, as recorded in Table 5.

Protein Content
As the data presented in Figure 11 indicate, U.f. polysacch. 5 mg/mL, U.f. polysacch. 10 mg/mL, and U.l. polysacch. 5 mg/mL induced the highest production of protein content, and U.l. polysacch. 10 mg/mL induced the lowest of the different selected plant seedlings of Zea mays; the protein contents were 96.76, 85.49, 74.54 and 69.75, respectively. These results follow those of Christobel [66] and Asir and Saravanababu [94], who pointed out that there was an increase in the protein content at the lower concentrations of seaweed extract-treated plants. In the present study, the observed significant increases in the amylase and protease activities of both tested plant seedlings in response to the effect of the priming agents are in good accord with the rise in growth, as well as the carbohydrate and protein composition. The results show that the low concentrations of both U.f. polysacch and U.l. polysacch are more effective with regard to growth parameters, antioxdant activities and protein contents; this may be due to the high concentrations of polysaccharide having more viscosity than the low concentrations. These results are comparable to those obtained by Siddhanat et al. [95], who reported that increasing the concentration of Ulva polysaccharide solutions lead to an increase in viscosity. The lower viscosity of chitosan improved the stability of the soil aggregates by 100 to 200 times after just one cycle of wetting-drying [96].

Protein Banding Pattern Pattern of Total Soluble Proteins in Zea mays Samples
The changes in the protein banding pattern in response to the priming of Zea plants with polysaccharide solutions are shown in Figure 12. The scanning of the gel indicates the existence of forty bands with molecular weights (MW) ranging from 8.921 to 258.772 Kilo Dalton (KDa) for all of the investigated samples. All of the studied samples showed complete identity in the number of bands. All of the treatments were characterized by eight common bands at molecular weights of 8.921, 17.854, 22.537, 29.326, 29.326, 35.19, 138.807, 170.837, and 258.772 KDa, respectively. Polysaccharide supplementation increased the relative protein concentrations; this may be due to gibberellin-prompted processes during seed germination [97]. 138.807, 170.837, and 258.772 KDa, respectively. Polysaccharide supplementation increased the relative protein concentrations; this may be due to gibberellin-prompted processes during seed germination [97].

Conclusions
It can be concluded that the soluble extracted polysaccharides of U. fasciata and U. lactuca are sources of bioactive compounds with potential applications in agriculture as they enhance the antioxidant activity and carbohydrate and protein contents of Zea mays. U. fasciata contains the worst bioactive material, rather than U. lactuca. The lower concentrations of soluble polysaccharide are more effective with regard to the growth parameters, carbohydrates, protein contents and antioxidant activates of Zea mays than higher concentrations. Polysaccharide supplementation increased the relative protein concentrations of Zea mays. The application of seaweeds' soluble polysaccharide extracts as biofertilizers is an eco-friendly approach for sustainable agriculture and the environment.

Conclusions
It can be concluded that the soluble extracted polysaccharides of U. fasciata and U. lactuca are sources of bioactive compounds with potential applications in agriculture as they enhance the antioxidant activity and carbohydrate and protein contents of Zea mays. U. fasciata contains the worst bioactive material, rather than U. lactuca. The lower concentrations of soluble polysaccharide are more effective with regard to the growth parameters, carbohydrates, protein contents and antioxidant activates of Zea mays than higher concentrations. Polysaccharide supplementation increased the relative protein concentrations of Zea mays. The application of seaweeds' soluble polysaccharide extracts as biofertilizers is an eco-friendly approach for sustainable agriculture and the environment.